The nef gene of human and simian immunodeficiency viruses (HIV-1, HIV-2, SIV) codes for a protein that is critical for the development of AIDS. Rhesus monkeys inoculated with pathogenic strains of SIV develop an AIDS-like disease and eventually die as a consequence. Remarkably, animals infected with an isogenic strain of SIV missing Nef remain healthy and acquire long-term immunity that protects them from subsequent challenge with pathogenic strains of SIV (containing Nef). Likewise, viruses defective in the nef gene have been isolated from some humans who are experiencing long-term non-progressive HIV-1 infection. Although the mechanism of Nef action is unknown, an intriguing discovery is that it binds tightly to SH3 domains of certain Src-family tyrosine kinases, which are proteins with diverse and critically important roles in many cellular signaling pathways. The crystal structure of the conserved core of HIV-1 Nef has been determined at 2.5 _ resolution in complex with the SH3 domain of a mutant Fyn tyrosine kinase (Arg 96 -> Ile), to which Nef binds tightly . Phases were determined to 3.3 _ using data measured at beamline X4A, BNL. Data measured to 2.5 _ at CHESS were used for refinement of the structure. The conserved "PxxP" sequence motif of Nef, known to be important for optimal viral replication, is part of a polyproline type II helix that engages the SH3 domain in a manner resembling closely the interaction of isolated peptides with SH3 domains. The Nef:SH3 structure also reveals how high affinity and specificity in the SH3 interaction is achieved by the presentation of the PxxP motif within the context of the folded structure of Nef. The design or discovery of compounds that bind to Nef may have potential therapeutic implications, and the structure of the complex that we have determined